GLYCOSYLATION ENHANCES MALONDIALDEHYDE BINDING TO PROTEINS

To determine whether glycosylation of proteins increases their suscept ibility to modification with malondialdehyde (MDA), bovine serum album in, which was pretreated with 500 mg/dl dextrose at 37 degrees C for 0 , 1, 2, and 4 weeks, were incubated with 100 mM MDA at 37 degrees C fo r 24 h. The MDA content of the protein samples were determined after d ialysis using thiobarbituric acid (TEA) assay. In addition, a specific anti-MDA protein antiserum was used to demonstrate MDA proteins with immunoblotting technique. The MDA content of BSA preincubated with dex trose for 4 weeks and reincubated with MDA (0.0649 +/- 0.0019 mu g MDA /mg protein) was significantly higher (p < .001) than the MDA content of BSA preincubated with dextrose for only one (0.0227 +/- 0.0031 mu g /mg) or two (0.0347 +/- 0.0034 mu g/mg weeks or the MDA content of non glysolated BSA incubated with MDA at the same experimental conditions (0.0201 +/- 0.0029 mu g/mg). These differences could also be found in the immunoblots. However, the correlation of TEA assay with the estima tes on immunoblots was poor. It is likely that the immune-blotting ass ay is more of an estimate of the number of BSA molecules modified with MDA, rather than MDA content of each BSA molecule. It is concluded th at in vitro glycosylation of proteins increases their susceptibility t o MDA-modification. This may well be an additional pathway of diabetes -related modification of proteins.